remaking the brain - University of Otago

REMAKING THE BRAIN
18
LISTENER OCTOBER 19 2013
THE
GREAT
GENE
QUEST
A scientist working on a shoestring
budget in a cramped Dunedin office
has taken a ground-breaking step
towards healing babies born with brain
damage. by REBECCA MACFIE photos by DAVID WHITE
I
Stephen Robertson:
a world-class win.
OCTOBER 19 2013 www.listener.co.nz
●
n the week that New Zealanders watched victory dissolve
into defeat on the waters of San
Francisco Bay, Professor Stephen
Robertson came to work as normal
in his cramped Dunedin laboratory, filled with a sense of quiet
satisfaction that his own small
team had once again achieved
a world-class win. There was
no ticker-tape parade or civic
ceremony to celebrate their
achievement. As Robertson stood in the
city’s main street at lunchtime, barely a soul
knew who he was, let alone that he had
achieved something that could profoundly
change lives.
The exhaustively peer-reviewed pages of
Nature Genetics represent some of the most
fiercely contested territory in the scientific world. It is the foremost international
journal for genetic research and on September 20, for the fourth time in a decade,
it revealed to the scientific community
ground-breaking work led by Robertson.
The latest paper goes by a dizzying title:
“Mutations in genes encoding the cadherin receptor-ligand pair DCHS1 and FAT4
disrupt cerebral cortical development”. Significantly, Robertson’s name appears at the
end of a list of 32 local and international collaborators, indicating that this family man
from Mosgiel was the intellectual leader of
the five-year research effort that yielded
what Melbourne paediatric neurologist Professor Ingrid Scheffer calls an “extensive,
meticulous and fascinating” body of work
that provides “seminal insights”.
Robertson’s group has made a pioneering discovery about two genes involved in
brain development, and how they influence
and regulate the way neural stem cells build
the human brain as the fetus grows in the
uterus. In 11 pages of dense, dry scientific
language, the Nature Genetics article provides
important clues as to how neural stem cells
might be harnessed and deployed to repair
the brains of babies damaged through the
likes of birth trauma, asphyxia, congenital
disease or even infant stroke (a condition
more frequent in newborns than previously
thought).
“This is a long-term goal, but understanding the biological pathways involved is the
essential first step,” says Scheffer, who is the
professor of paediatric neurology at the University of Melbourne. “The aim would be
to mobilise these special cells [neural stem
cells] from within the infant brain to repair
the lost cells.”
19
REMAKING THE BRAIN
ORCHESTRATING DEVELOPMENT
Robertson knows to keep the language sober
and the expectations realistic, but he shares
this startlingly ambitious goal. “The hope is
that if we can understand some of the ‘language’ going on in the infant brain between
stem cells, particularly in the context of an
infant brain that has become damaged, perhaps we can leap in and encourage the brain
to repair itself.”
The package of knowledge behind all
this is excruciatingly complex, but Robertson – the Cure Kids Professor of Paediatric
Genetics at the University of Otago – is
an expert at translating it. “The human
brain develops from a single layer of cells
that forms very early at the top end of the
developing embryo. This layer of cells, called
the neuroepithelium, houses a population
of stem cells that have two major developmental tasks to perform to orchestrate brain
development.
“The first is that they must renew themselves. But secondly, and crucially, they must
also retain the capability to turn into any
type of brain cell when required. As the
brain develops, these two tasks need to be
regulated: on one hand, the cells need to proliferate, and on the other, at appropriately
timed points during brain development,
they must change into mature brain cells
– neurons and the like – to actually make a
functioning and integrated brain.”
There is much excitement in the research
world about the possibility of growing adult
stem cells and injecting them into the diseased brains of Alzheimer’s or Parkinson’s
sufferers. But Robertson says the potential
for intervention in infant brains is possibly
far greater.
“We know that infants are born with this
very substantial population of neural stem
cells that are still very active and still migrating and doing their stuff to form the brain.
The infant brain is very plastic; the adult
brain is much less so.”
Until now, little has been known about
what prompts neural stem cells to take up
their mature position and function in the
developing brain. In a triumph of collaboration between Robertson’s tight team of
PhD students and post-doctoral researchers
in Dunedin and some of the world’s most
respected neuroscientists – along with helpers from Canada, France, the Netherlands
and the UK – something of that mystery
has been unlocked. In essence, they have
discovered a “radio signal” used by neural
stem cells to communicate with each other
as they conduct brain development.
“This is genuinely great science,” says
Professor Russell Snell, of the University
20
“This is genuinely
great science … It’s
a beautiful story.”
of Auckland’s neurogenetics group. “It’s a
beautiful story that starts with basic clinical science. The thing that makes Stephen
Robertson unique is that he [is] a geneticist
who sees patients with rare disorders, and
he is also a research scientist. So he’s not
just a geek, but he is a geek as well. He has
an incredibly strong drive to do good things
for his patients.”
INTERNATIONAL NETWORK
Robertson’s latest journey of discovery began
in about 2007, on the back of his rising
international reputation as a clinical and
research geneticist (of which more later).
Through his networks with other clinicians
around the world who see patients with
genetic disorders, his lab had accumulated
a bank of 3000 vials of DNA from people
born with rare conditions. As he met up
with his peers at international meetings and
kept in contact via email, it became clear
that a new and extremely unusual genetic
syndrome was being identified. The sufferers
of the condition had a raft of symptoms:
clawed hands, widely spaced eyes, tiny ears
and deafness, intellectual disability – and
a large band of “grey” matter stuck in the
middle of the brain, rather than forming
the outermost layer on the cerebral cortex
where it is supposed to be.
Robertson had already seen this abnormality in the construction of the brain in
earlier work, and it was known to give rise
to a seizure disorder in females. Yet here it
was again, presenting in these patients with
a completely different clinical syndrome.
For some reason the neural stem cells in
the developing brain of these children had
failed to migrate to their correct position
and take on their specialist roles.
The parents of children born with this
mysterious condition didn’t even have
the benefit of a clear diagnosis or a name
to describe it (although in the past couple
of years it has become known as Van
Maldergem syndrome, after the Belgian
geneticist who described the first case). At
the time, Robertson had working in his
Dunedin lab a talented young PhD student
from Horowhenua, Mary Gray. In 2008, she
set to work studying this rare syndrome, and
quickly deduced that the gene responsible
for the condition must lie on chromosome
11. But which of thousands might it be?
“I remember her coming into the office
and saying to me, ‘This is where it’s got to
be.’ But, wow, it was like looking at an aerial
photograph and trying to spot a single letter
box,” recalls Robertson.
He knew what a slow and painstaking
task it would be to track down the culprit
gene the old-fashioned way. He’d done it
himself in 2002 when, as a PhD student at
Oxford, he’d spent three years searching for
the gene responsible for the tragic malformation and death of seven babies born to a
West Auckland family. But by 2008, genetic
research technology had advanced dramatically, massively speeding up the process of
identifying the genes and mutations implicated in congenital disorders.
So Gray and Robertson opted to use what’s
called Massively Parallel Genomic Sequencing technology, which meant sending the
DNA of chromosome 11 to a specialist company in Connecticut, where it was shredded
and rinsed clean of the 98% of DNA that
does not code for proteins (the stuff that was
once referred to as “junk DNA” but is now
known to be functional), and then decoded.
A few weeks later the information was
mailed back on a couple of CDs to Robertson’s Hanover St lab. Gray went to work,
hunting through the lines of code for
the genetic “spelling mistakes” that were
causing the syndrome. Before long, she
implicated the gene responsible, identifiable by mutations that were repeated among
three unrelated families with the syndrome.
A STROKE OF LUCK
This alone would have been worthy of a
hearty celebration and publication in a
scholarly journal. But Robertson and Gray
sensed there was much more to learn; they
wanted to know not only what had happened in the culprit gene to cause this
condition but how.
The gene coded for a protein named
Dachsous1. Robertson says he “didn’t know
it from a bar of soap” at the time. He had
always been focused on a set of genes known
as filamins, so getting to grips with Dachsous required masses of study to find out
what was already known about it.
By a great stroke of luck, it turned out that
it had been intensively studied in fruit flies,
but “jumping from what is known about
this gene in flies to understanding its role in
a mammal, let alone a human, is a big, big
leap. That was a mind-blowing moment.”
What the fly geneticists had discovered
was that the Dachsous1 gene sits embedded in the membrane on the outside of cells
LISTENER OCTOBER 19 2013
Clockwise from left:
Tania Gunn; Francis
Collins; Robertson
in his office;
Magdalena Götz,
Andrew Wilkie;
Robertson and
family on holiday in
Wales in 2011.
OCTOBER 19 2013 www.listener.co.nz
work looking for misspellings on the FAT4
gene. And there they were: the DNA of the
three remaining unrelated families with Van
Maldergem syndrome all had mutations distributed throughout the gene.
By now, Robertson sensed that they
were onto something more than just the
cause of a rare congenital syndrome and
the underpinning role of the two culprit
genes. He went back to the literature on
fly biology, and found that the interaction
between FAT4 and Dachsous1 was known
to produce an instruction – something akin
to a language or a radio signal – that was
involved in the development of the embryo
of all multicellular organisms. There was
already considerable knowledge about that
signal, which went by the peculiar name
of “Hippo signalling” and had long been
thought to be involved in regulating the size
of internal organs such as the kidney and
liver. But what Robertson and Gray didn’t
know was what it had to do with the formation of the brain.
Then, in another stroke of luck, a Japanese paper published in 2009 implicated
21
DAVID WHITE; GETTY IMAGES; PROF STEPHEN ROBERTSON
and binds onto a gene that codes for a protein called FAT4 – encoded by the “biggest,
ugliest gene you’ve ever seen” – located on
the surface of an adjacent cell. Robertson
figured both Dachsous1 and FAT4 were playing a role in the cause of Van Maldergem
syndrome, but looking for the mutation
on FAT4 that might be causing the trouble
was going to be like “looking for a bolt out
of place on the Starship Enterprise”.
Once again, they sent a batch of DNA off
for sequencing, this time to a collaborator in
London. When it came back, Gray went to
REMAKING THE BRAIN
Dachsous1 and FAT4 in the developing
brains of mice. It appeared that if either of
the two genes was damaged or disabled, the
expression of the other became defective.
Smack in the centre of the tiny brain of an
embryonic mouse, the two genes were playing a commanding role in the way neural
stem cells did their work.
Robertson and his team knew the two
genes were implicated in Van Maldergem
syndrome, and now they knew they also
played a key role in the development of
a mouse brain. But how to connect these
fragments of knowledge and give them
meaning for human health?
BEAUTIFUL TOOLS
It was time to search for a collaborator in
what Robertson likens to a quest to figure
out the “electrical engineering” of neural
stem cells: “I had this feeling that we were
onto an understanding of a human disorder
that tells us something about what channels these neural stem cells are tuned into
and what language they use to regulate
themselves.”
He scoured the literature and settled on
a German neuroscientist, Magdalena Götz,
internationally recognised for her work in
discovering the nature of neural stem cells
and the leader of a large research group.
Götz had developed “beautiful tools where
she can interfere with genes in the brains of
embryonic mice in a very precise way – to
disable genes and then observe the effect
of this manipulation on their subsequent
function and behaviour. This is in real time,
in a live mouse embryo.”
Robertson emailed her with news that his
small Dunedin lab had identified the role of
two genes in a rare human condition, and
asked whether she was keen to collaborate.
Götz responded immediately, and a powerful research partnership began. Götz and
her senior research fellow, Sylvia Cappello,
would inject a fluorescent protein into the
embryonic brain of mice, alongside a chemical that disabled either Dachsous1 or FAT4,
then put the embryo back into the uterus
and watch how the intervention affected
the behaviour of the fluorescent stem cells.
In this way, they were able to replicate
Van Maldergem syndrome in mice. Cappello then went further, observing the “radio
signal” that seemed to be orchestrating the
damage, and showing that by interfering at
the right time with that signal, the damage
to the brain could be corrected.
“It was a heady moment to realise that on
one hand we could produce a developmental disorder within the brain of a mammal
with one intervention, and then with the
22
“It does show that
it is possible to lead
these international
collaborations from
New Zealand.”
other, reverse it,” says Robertson. These
experiments provided proof that they had
found a mode of communication used by
these two key genes to regulate the work of
stem cells in the developing brain.
Götz says her lab could not have made
this breakthrough without the benefit of
Robertson’s new knowledge of the effect of
the two genes on human development; Robertson says his lab could not have modelled
the effect in mice so quickly and precisely
without Götz’s techniques.
“It does show that it is possible to lead
these international collaborations from New
Zealand,” he says.
CURING KIDS
As Russell Snell points out, Robertson’s latest
research achievement is not a one-off. “He
regularly does this sort of work, and he does
it on a shoestring. Yes, he has funding, but
nowhere near the funding of the large US
and European labs.”
For Robertson, it all began as a young
paediatric registrar at Starship Children’s
Hospital in the 1990s. Having grown up in
a large Catholic family in Hawke’s Bay, he
had gone to the University of Otago to study
medicine and graduated with distinction,
winning the Prince of Wales Prize for the
most outstanding student completing an
undergraduate degree, and coming top of
his medical school class.
Genetics wasn’t even taught at the university while he was training, and only a small
number of genetic disorders had been solved
at the molecular level – the race to find the
gene for cystic fibrosis was won in the late
1980s by an American group led by Francis
Collins while Robertson was a student. But
it was a field of research that was about to
undergo explosive development.
The idea of curing children had enormous
appeal, and in his second year out of medical school he went to the newly opened
Starship as a house surgeon before being
fast-tracked into a registrar role.
Before long, he met the family who would
change his life and launch his career as a
world-class geneticist.
June Miru and her family had come into
the care of Starship paediatrician Tania
Gunn in 1989. The close-knit whanau had
begun to fear that they were afflicted by a
makutu – a curse. June, the matriarch, had
lost her first-born boy in 1963 – a child so
malformed that she wasn’t permitted to see
him. She went on to give birth to another
son and four daughters – all healthy. Then
in 1988, her daughters began having babies.
And the babies, if they were boys, were
mostly dying at birth, their tiny little bodies
cruelly twisted and tangled, with lungs compressed within malformed rib cages, with
defective hearts and, in some cases, with
their abdomens open and their intestines
on the outside.
Gunn had taken the Miru family under
her wing and, along with Professor David
Becroft and the up-and-coming registrar Stephen Robertson, wrote a paper describing
the syndrome.
The cause was clearly genetic and passed
on by the mother. Robertson wrote to an
eminent geneticist at Oxford, Andrew
Wilkie, and asked nervously if he would
help him find the answer to the Miru family’s torment. Wilkie made a start, but it soon
became clear to Robertson that he didn’t
have much time to invest in solving the
mystery. Instead, Wilkie suggested to Robertson that he come to Oxford and have a
go at finding the gene himself. “I thought,
‘Wow! Going to Oxford, finding a disease
gene!’ – it pressed all my buttons.”
But Robertson had no training in genetics. Everything he knew was self-taught.
If he was to crack the answer for the Miru
family, he needed to be properly schooled in
the discipline. So, with his wife, GP Robyn
Blake, and their infant son Nick in tow, he
went to the Murdoch Institute in Melbourne
– an eminent outfit bankrolled by Rupert
Murdoch’s philanthropic mother, Dame
Elizabeth Murdoch – and studied genetics
as a sub-specialty for three years.
Wilkie continued to chip away at the
Miru family’s problem in the meantime,
but progress was slow. Again, Wilkie put it
to Robertson that he should come to Oxford
and hunt for the gene.
OFF TO OXFORD
So, funded by a Nuffield Scholarship, he,
Robyn and their growing family (by then
Nick had a baby brother, Mark), headed to
the UK in 1999 with the objective of getting
an Oxford PhD in genetics and delivering
an explanation to June Miru and her family.
“I was absolutely obsessed,” recalls Robertson of his intensive three years there. “In
some ways it felt as if I was looking under
every stone in a river bed. It didn’t feel
LISTENER OCTOBER 19 2013
June Miru and
her daughter
Noki-Jane Miru
Saunders; the
wider Miru
whanau, pictured
below in 2003,
lost seven babies
as the result of a
gene mutation.
OCTOBER 19 2013 www.listener.co.nz
– Robertson began looking at a gene he’d
previously ignored. In 1998, a mutation in
a gene that codes for a protein called filamin
A had been discovered to be the cause of
a syndrome characterised by epileptic seizures. Robertson had initially discounted it,
thinking that if it caused seizures it would
have nothing to do with the complex skeletal syndrome suffered by the Miru family.
Using the laborious techniques available
at the time, he noticed a suspicious-looking
aberration in this gene in one of the sufferers of the syndrome, but he was nervous
about believing what he was seeing. One
evening, while having a pint with his laboratory colleagues, he grabbed a copy of the
Salvation Army magazine Watchtower that
was lying on a table, jotted his findings
23
DAVID WHITE; JANE USSHER
terribly creative at times and sometimes it
felt quite desperate.”
His hunting ground was two million base
pairs of DNA, and within that, about 60
genes. He scoured the world and found
other families suffering the same disorder as the Mirus; most had had only one
baby with the syndrome and some, frozen
with fear, had no more. What made the
Mirus unique was that they had continued
to have babies, providing a rich statistical foundation from which to search for
genetic clues.
There were false starts when he would go
home and tell Robyn that he thought he’d
found the responsible gene, only to discover he was wrong. He remained in close
contact with June Miru and the family,
phoning regularly and keeping them up to
date – even when he felt as if he was banging his head against a brick wall. While
his hunt for the offending gene continued,
June’s youngest daughter, Noki-Jane, lost
her second baby boy to the syndrome.
But June says she never doubted that this
“young pup”, whom she came regard as a
son, would deliver the answer. “He knew we
were sitting waiting.”
Eventually – and increasingly desperate as
his three-year scholarship was running out
REMAKING THE BRAIN
down in a margin and asked the others if
they “smelt a rat”.
“They looked back at me over the top
of their lagers and said, ‘Oh, you doofus!
You’ve found the bloody thing.’”
Robertson’s findings – co-authored by
Andrew Wilkie – were published in 2003 in
Nature Genetics. More importantly, he was
able to call the Miru family together and
give them the gift of an explanation, a name
for the gene responsible (FLNA), and the
ability to test the women of the families to
ascertain if they were carriers of the mutation and to test their unborn babies to find
out if they were affected.
Not long after, the family presented him
with a small trophy that he still keeps on his
desk in his tiny Dunedin office. The inscription reads: “The What Took You So Long
Award” – a dry and understated token of
gratitude for the dry and understated man
who came into their lives and lightened
their misery. “He lifted a veil for us as a
family,” says June Miru.
The inscription on the
small trophy reads:
“The What Took You
So Long Award.”
collaboration with American scientists, they
discovered a mutation in another filamincoding gene (FLNB) that caused children
to be born with their joints dislocated from
their sockets. The work was published in
Nature Genetics in 2004.
Clinical geneticists from around the world
began asking him to enrol their patients in
his research programme, and rapidly the
fridges in the Dunedin lab were filled with
thousands of DNA samples.
At the same time, Cure Kids decided not
only to part-fund the Otago chair in paediatric genetics (one of three research chairs
ROBERTSON FAMILY ARCHIVE
FARREACHING IMPLICATIONS
Sadly, Tania Gunn, who had initially taken
the family into her care, died of cancer
just months before Robertson made his
breakthrough.
The implications of the discovery went far
beyond the Miru family and others with the
same syndrome. The offending gene turned
out to be a “many-faced beast, implicated
in a whole raft of congenital syndromes”.
Where the filamin proteins had previously
been thought to play a rather minor role in
cell development, Robertson’s disclosures
contributed to showing that they were a
crucial regulator. The discovery launched
Robertson’s research career on two parallel
paths: learning more about the gene’s role in
skeletal malformations and in neurological
abnormalities.
In the middle of all this, he was asked to
come home to New Zealand and establish
a new research facility in Dunedin, jointly
funded by Cure Kids and the University of
Otago. Still only in his mid-thirties and still
polishing off his PhD, he was being asked
to cold-start a genetics research lab on the
opposite side of the planet from Oxford. “It
all felt a bit premature,” he recalls.
But he and Robyn were keen to raise the
family (which by then included Isabelle,
born in Oxford) in a smaller centre, and
the decision was made.
Even before he got the new Dunedin lab
properly set up, he made another major
breakthrough. He and research technician Tim Morgan were working with
borrowed and shared equipment when, in a
24
Robertson (at rear) and his research team,
with Mary Gray (in pink). Others, from left, are
Sarah Holman, Tim Morgan, Zandra Jenkins,
Dee Yang, Phil Daniel, Sarah Cardoso, Sophia
Cameron-Christie and Heather Tiffin.
funded by the charity), but to establish
an “enablement” fund of $1 million over
five years that can be spent on blue-skies
research. Most New Zealand scientists have
to scrabble around for funding that is often
short-term but, Robertson says, the stability of the Cure Kids funding gives him and
his colleagues space to think and “tinker”.
“They understand that science works by fiddling around in your tool shed, kicking the
tyres on ideas, trying things out … There is
so little provision for that in New Zealand.”
In 2009, his group headlined in Nature
Genetics again, this time with research that
disrupted contemporary thinking about the
role of a mutation in what was known as
a “cancer gene”, called WTX, which was
thought to predispose carriers to the childhood cancer Wilms tumour. Robertson’s
work showed that being born with a WTX
mutation resulted in a severe bone disorder,
rather than cancer.
The work won him the Health Research
Council’s Liley Medal in 2010. By this time
there was rising international confidence
that his small genetics lab at the bottom of
the world could deliver, and the work on
Van Maldergem syndrome that would eventually lead to the breakthrough published in
Nature Genetics last month was under way.
THE TYRANNY OF DISTANCE
When Robertson left Oxford in 2003, his
mentor Andrew Wilkie told him that in
moving back to distant New Zealand he was
electing to make his research career 50%
more difficult. A fair comment? Robertson
adopts his characteristically quizzical
expression, head tilted a little to the side,
and acknowledges that Wilkie might have
been exaggerating a little but was possibly
right. Despite email and the internet, there is
still the tyranny of distance and the absence
of a large local community of
research peers. “I can’t just go
out into the corridor and find a
Magdalena [Götz].”
But there are compensations.
What his lab lacks in scale
it makes up for in close and
productive relationships – both
within the university and with
the members of the public
who financially support his
research through Cure Kids.
New Zealanders are generous
and supportive, he says, and
they are “just one step away” – he spends
large amounts of time explaining his work
to groups.
There’s also the lifestyle. On Robertson’s
computer screen is a glorious photo of
the bright red Brewster Hut, set on a high
tussock-clad ridge near Haast where he
tramped recently. A few days after our
interview, he and the children were off
to Queenstown to support Robyn while
she competed in the Spring Challenge
multisport event. And the following week
he was in one of his regular clinical sessions
seeing children with rare genetic disorders
– the coal-face work that takes up 30% of
his time and keeps him constantly in touch
with the real purpose of his geeky scientific
endeavours.
Whether he’s in Dunedin or Oxford, that,
in the end, is what matters to Robertson.
“Working with children is something I find
enormously rewarding – knowing that we
can actually make a difference to a life that
is unfolding.” l
LISTENER OCTOBER 19 2013